BRG1-associated Factor Complex Research Articles

Alpha-synuclein promotes PRMT5-mediated H4R3me2s histone methylation by interacting with the BAF complex.

α-Synuclein (αS) is a key molecule in the pathomechanism of Parkinson's disease. Most studies on αS to date have focused on its function in the neuronal cytosol, but its action in the nucleus has also been postulated. Indeed, several lines of evidence indicate that overexpressed αS leads to epigenomic alterations. To clarify the functional role of αS in the nucleus and its pathological significance, HEK293 cells constitutively expressing αS were used to screen for nuclear proteins that interact with αS by nanoscale liquid chromatography/tandem mass spectrometry. Interactome analysis of the 229 identified nuclear proteins revealed that αS interacts with the BRG1-associated factor (BAF) complex, a family of multi-subunit chromatin remodelers important for neurodevelopment, and protein arginine methyltransferase 5 (PRMT5). Subsequent transcriptomic analysis also suggested a functional link between αS and the BAF complex. Based on these results, we analyzed the effect of αS overexpression on the BAF complex in neuronally differentiated SH-SY5Y cells and found that induction of αS disturbed the BAF maturation process, leading to a global increase in symmetric demethylation of histone H4 on arginine 3 (H4R3me2s) via enhanced BAF-PRMT5 interaction. Chromatin immunoprecipitation sequencing confirmed accumulated H4R3me2s methylation near the transcription start site of the neuronal cell adhesion molecule (NRCAM) gene, which has roles during neuronal differentiation. Transcriptional analyses confirmed the negative regulation of NRCAM by αS and PRMT5, which was reconfirmed by multiple datasets in the Gene Expression Omnibus (GEO) database. Taken together, these findings suggest that the enhanced binding of αS to the BAF complex and PRMT5 may cooperatively affect the neuronal differentiation process.

Structural disruption of BAF chromatin remodeller impairs neuroblastoma metastasis by reverting an invasiveness epigenomic program

BackgroundEpigenetic programming during development is essential for determining cell lineages, and alterations in this programming contribute to the initiation of embryonal tumour development. In neuroblastoma, neural crest progenitors block their course of natural differentiation into sympathoadrenergic cells, leading to the development of aggressive and metastatic paediatric cancer. Research of the epigenetic regulators responsible for oncogenic epigenomic networks is crucial for developing new epigenetic-based therapies against these tumours. Mammalian switch/sucrose non-fermenting (mSWI/SNF) ATP-dependent chromatin remodelling complexes act genome-wide translating epigenetic signals into open chromatin states. The present study aimed to understand the contribution of mSWI/SNF to the oncogenic epigenomes of neuroblastoma and its potential as a therapeutic target.MethodsFunctional characterisation of the mSWI/SNF complexes was performed in neuroblastoma cells using proteomic approaches, loss-of-function experiments, transcriptome and chromatin accessibility analyses, and in vitro and in vivo assays.ResultsNeuroblastoma cells contain three main mSWI/SNF subtypes, but only BRG1-associated factor (BAF) complex disruption through silencing of its key structural subunits, ARID1A and ARID1B, impairs cell proliferation by promoting cell cycle blockade. Genome-wide chromatin remodelling and transcriptomic analyses revealed that BAF disruption results in the epigenetic repression of an extensive invasiveness-related expression program involving integrins, cadherins, and key mesenchymal regulators, thereby reducing adhesion to the extracellular matrix and the subsequent invasion in vitro and drastically inhibiting the initiation and growth of neuroblastoma metastasis in vivo.ConclusionsWe report a novel ATPase-independent role for the BAF complex in maintaining an epigenomic program that allows neuroblastoma invasiveness and metastasis, urging for the development of new BAF pharmacological structural disruptors for therapeutic exploitation in metastatic neuroblastoma.

Expanding the phenotype associated with SMARCC2 variants: a fetus with tetralogy of Fallot

BackgroundCoffin-Siris syndrome-8 (CSS8) is a rare autosomal dominant disorder caused by variants in SMARCC2, a core subunit of the chromatin-remodeling complex BRG1-associated factor (BAF). The clinical characteristics of this disorder have not been entirely determined because of the rarity of clinical reports. The BAF complex plays a crucial role in embryogenesis and cardiac development, and pathogenic variants in genes encoding the components of the BAF complex have been associated with congenital heart disease (CHD). However, variants in SMARCC2 have not been reported in patients with CHD.Case presentationA 28-year-old primigravida was referred at 24 weeks gestation for prenatal echocardiography. The echocardiographic findings were consistent with a prenatal ultrasound diagnosis of tetralogy of Fallot (TOF). After detailed counseling, the couple decided to terminate the pregnancy and undergo genetic testing. A trio (fetus and the parents) whole-exome sequencing (WES) and copy number variation sequencing (CNV-seq) were performed. CNV-seq identified no chromosomal abnormalities. WES analysis revealed a pathogenic, de novo heterozygous frameshift variant in SMARCC2 (NM_003075.5: c.3561del, p.Leu1188fs). The genetic diagnosis of CSS8 was considered given the identification of the SMARCC2 pathogenic variant.ConclusionsWe report the first prenatal case with the SMARCC2 variant. The presence of CHD further broadens the phenotypic spectrum of SMARCC2-related disease.

Insights Into the Emerging Role of Baf53b in Autism Spectrum Disorder.

Accurate and precise regulation of gene expression is necessary to ensure proper brain development and plasticity across the lifespan. As an ATP-dependent chromatin-remodeling complex, the BAF (Brg1 Associated Factor) complex can alter histone-DNA interactions, facilitating dynamic changes in gene expression by controlling DNA accessibility to the transcriptional machinery. Mutations in 12 of the potential 29 subunit genes that compose the BAF nucleosome remodeling complex have been identified in several developmental disorders including Autism spectrum disorders (ASD) and intellectual disability. A novel, neuronal version of BAF (nBAF) has emerged as promising candidate in the development of ASD as its expression is tied to neuron differentiation and it’s hypothesized to coordinate expression of synaptic genes across brain development. Recently, mutations in BAF53B, one of the neuron specific subunits of the nBAF complex, have been identified in patients with ASD and Developmental and epileptic encephalopathy-76 (DEE76), indicating BAF53B is essential for proper brain development. Recent work in cultured neurons derived from patients with BAF53B mutations suggests links between loss of nBAF function and neuronal dendritic spine formation. Deletion of one or both copies of mouse Baf53b disrupts dendritic spine development, alters actin dynamics and results in fewer synapses in vitro. In the mouse, heterozygous loss of Baf53b severely impacts synaptic plasticity and long-term memory that is reversible with reintroduction of Baf53b or manipulations of the synaptic plasticity machinery. Furthermore, surviving Baf53b-null mice display ASD-related behaviors, including social impairments and repetitive behaviors. This review summarizes the emerging evidence linking deleterious variants of BAF53B identified in human neurodevelopmental disorders to abnormal transcriptional regulation that produces aberrant synapse development and behavior.

Expanding the Spectrum of BAF-Related Disorders: De Novo Variants in SMARCC2 Cause a Syndrome with Intellectual Disability and Developmental Delay

SMARCC2 (BAF170) is one of the invariable core subunits of the ATP-dependent chromatin remodeling BAF (BRG1-associated factor) complex and plays a crucial role in embryogenesis and corticogenesis. Pathogenic variants in genes encoding other components of the BAF complex have been associated with intellectual disability syndromes. Despite its significant biological role, variants in SMARCC2 have not been directly associated with human disease previously. Using whole-exome sequencing and a web-based gene-matching program, we identified 15 individuals with variable degrees of neurodevelopmental delay and growth retardation harboring one of 13 heterozygous variants in SMARCC2, most of them novel and proven de novo. The clinical presentation overlaps with intellectual disability syndromes associated with other BAF subunits, such as Coffin-Siris and Nicolaides-Baraitser syndromes and includes prominent speech impairment, hypotonia, feeding difficulties, behavioral abnormalities, and dysmorphic features such as hypertrichosis, thick eyebrows, thin upper lip vermilion, and upturned nose. Nine out of the fifteen individuals harbor variants in the highly conserved SMARCC2 DNA-interacting domains (SANT and SWIRM) and present with a more severe phenotype. Two of these individuals present cardiac abnormalities. Transcriptomic analysis of fibroblasts from affected individuals highlights a group of differentially expressed genes with possible roles in regulation of neuronal development and function, namely H19, SCRG1, RELN, and CACNB4. Our findings suggest a novel SMARCC2-related syndrome that overlaps with neurodevelopmental disorders associated with variants in BAF-complex subunits.

A non-canonical BRD9-containing BAF chromatin remodeling complex regulates naive pluripotency in mouse embryonic stem cells

The role of individual subunits in the targeting and function of the mammalian BRG1-associated factors (BAF) complex in embryonic stem cell (ESC) pluripotency maintenance has not yet been elucidated. Here we find that the Bromodomain containing protein 9 (BRD9) and Glioma tumor suppressor candidate region gene 1 (GLTSCR1) or its paralog GLTSCR1-like (GLTSCR1L) define a smaller, non-canonical BAF complex (GBAF complex) in mouse ESCs that is distinct from the canonical ESC BAF complex (esBAF). GBAF and esBAF complexes are targeted to different genomic features, with GBAF co-localizing with key regulators of naive pluripotency, which is consistent with its specific function in maintaining naive pluripotency gene expression. BRD9 interacts with BRD4 in a bromodomain-dependent fashion, which leads to the recruitment of GBAF complexes to chromatin, explaining the functional similarity between these epigenetic regulators. Together, our results highlight the biological importance of BAF complex heterogeneity in maintaining the transcriptional network of pluripotency.

First data from a parent-reported registry of 81 individuals with Coffin-Siris syndrome: Natural history and management recommendations.

Coffin-Siris syndrome (CSS; MIM 135900) is a multisystem congenital anomaly syndrome caused by mutations in the genes in the Brg-1 associated factors (BAF) complex. Classically, individuals with CSS have been described with hypo- or aplasia of the fifth digit nails or phalanges (hence the term "fifth digit syndrome"). Other physical features seen include growth restriction, coarse facial features, hypertrichosis or hirsutism, sparse scalp hair, dental anomalies, and other organ-system abnormalities. Varying degrees of developmental and intellectual delay are universal. To date, approximately 200 individuals have been described in the literature. With the advent of large-scale genetic testing such as whole-exome sequencing is becoming more available, more individuals are being found to have mutations in this pathway, and the phenotypic spectrum appears to be broadening. We report here a large cohort of 81 individuals with the diagnosis of CSS from the first parent-reported CSS/BAF complex registry in an effort to describe this variation among individuals, the natural history of the syndrome, and draw some gene-phenotype correlations. We propose that changes in the BAF complex may represent a spectrum of disorders, including both ARID1B-related nonsyndromic intellectual disability (ARID1B-ID) and CSS with classic physical features. In addition, we offer surveillance and management recommendations based on the medical issues encountered in this cohort to help guide physicians and patients' families.

Heterozygous variants in ACTL6A, encoding a component of the BAF complex, are associated with intellectual disability.

Pathogenic variants in genes encoding components of the BRG1-associated factor (BAF) chromatin remodeling complex have been associated with intellectual disability syndromes. We identified heterozygous, novel variants in ACTL6A, a gene encoding a component of the BAF complex, in three subjects with varying degrees of intellectual disability. Two subjects have missense variants affecting highly conserved amino acid residues within the actin-like domain. Missense mutations in the homologous region in yeast actin were previously reported to be dominant lethal and were associated with impaired binding of the human ACTL6A to β-actin and BRG1. A third subject has a splicing variant that creates an in-frame deletion. Our findings suggest that the variants identified in our subjects may have a deleterious effect on the function of the protein by disturbing the integrity of the BAF complex. Thus, ACTL6A gene mutation analysis should be considered in patients with intellectual disability, learning disabilities, or developmental language disorder.

Swi/Snf chromatin remodeling/tumor suppressor complex establishes nucleosome occupancy at target promoters

Precise nucleosome-positioning patterns at promoters are thought to be crucial for faithful transcriptional regulation. However, the mechanisms by which these patterns are established, are dynamically maintained, and subsequently contribute to transcriptional control are poorly understood. The switch/sucrose non-fermentable chromatin remodeling complex, also known as the Brg1 associated factors complex, is a master developmental regulator and tumor suppressor capable of mobilizing nucleosomes in biochemical assays. However, its role in establishing the nucleosome landscape in vivo is unclear. Here we have inactivated Snf5 and Brg1, core subunits of the mammalian Swi/Snf complex, to evaluate their effects on chromatin structure and transcription levels genomewide. We find that inactivation of either subunit leads to disruptions of specific nucleosome patterning combined with a loss of overall nucleosome occupancy at a large number of promoters, regardless of their association with CpG islands. These rearrangements are accompanied by gene expression changes that promote cell proliferation. Collectively, these findings define a direct relationship between chromatin-remodeling complexes, chromatin structure, and transcriptional regulation.

Target genes of the largest human SWI/SNF complex subunit control cell growth

The largest subunit of the mammalian SWI/SNF-A or BAF (BRG1-associated factor) chromatin-remodelling complex is encoded by two related cDNAs hOsa1/BAF250a and hOsa2/BAF250b that are unique to the BAF complex and absent in the related PBAF (Polybromo BAF). hOsa/BAF250 has been shown to interact with transcriptional activators and bind to DNA suggesting that it acts to target the remodelling complex to chromatin. To better understand the functions of hOsa2, we established inducible stable HeLa cell lines over-expressing FLAG-hOsa2 or a derivative lacking the ARID (AT-rich interactive domain) DNA-binding domain. Immunopurification of complexes containing hOsa2 that was followed by mass spectrometry and immunoblotting demonstrated the presence of BRG1 and known BAFs, but not hOsa1 or hBRM. Deletion of the ARID did not compromise the integrity of the complex. Induction of hOsa2 expression caused impaired cell growth and accumulation of cells in the G0/G1 cell cycle phase. Elevated levels of the p53 and p21 proteins were detected in these cells while c-Myc mRNA and protein levels were found to decrease. Chromatin immunoprecipitation and reporter assays suggested that hOsa2 had a direct effect on c-myc and p21 promoter activity. Thus hOsa2 plays an important role in controlling genes regulating the cell cycle.

Reconstitution of glucocorticoid receptor-dependent transcription in vivo.

We developed a model system to study glucocorticoid receptor (GR)-mediated chromatin remodeling by the BRG1 complex. Introduction of the BRG1 ATPase into the SW-13 cell line initiates the formation of a functional remodeling complex. This complex is able to induce transcriptional activation from a transiently transfected promoter with wild-type and chromatin-remodeling-deficient BRG1 mutants, suggesting that the complex possesses a coactivator function independent from remodeling. Transactivation from a chromatin template requires the BRG1 remodeling function, which induces regions of hypersensitivity and transcription factor loading onto the integrated MMTV promoter. We report that BRG1 remodeling activity is required for GR-mediated transactivation and that this activity cannot be replaced by other ATP-dependent remodeling proteins. Further characterization of the BRG1-associated factors (BAFs) present in these cells (for example, the expression of BAF250 but not BAF180) reveals that the BAF complex rather than the polybromo-associated BAF complex is the necessary and sufficient chromatin-remodeling component with which the receptor functions in vivo. These results in conjunction with previous findings demonstrate that the GR functions with multiple forms of the SWI/SNF complex in vivo.